Expanding the phenotype of Seckel syndrome associated with biallelic loss-of-function variants in CEP63

Seckel syndrome is an ultrarare autosomal recessive genetically heterogenous condition characterized by intrauterine and postnatal growth restriction, proportionate severe short stature, severe microcephaly, intellectual disability, and distinctive facial features including a prominent nose. Up to now, 40 patients with molecularly confirmed Seckel syndrome have been reported with biallelic variants in nine genes: ATR, CENPJ, CEP63, CEP152, DNA2, NIN, NSMCE2, RBBP8, and TRAIP. Homozygosity for nonsense variant (c.129G>A, p.43*) in CEP63 was described in three cousins with microcephaly, short stature, mild to moderate intellectual disability and diagnoses of Seckel syndrome. Here, we report a second family with three siblings who are compound heterozygous for loss-of-function variants in CEP63, c.1125T>G, p.(Tyr375*) and c.595del, p.(Glu199Asnfs*11). All siblings present with microcephaly, prominent nose, and intellectual disability but only one has severe short stature. Two siblings have aggressive behavior, a feature previously not reported in Seckel syndrome. This report adds two novel truncating variants in CEP63 and extends the clinical knowledge on CEP63-related conditions.     

Compound heterozygosity for loss‐of‐function GARS variants results in a multisystem developmental syndrome that includes severe growth retardation

Aminoacyl‐tRNA synthetases (ARSs) are ubiquitously expressed enzymes that ligate amino acids onto tRNA molecules. Genes encoding ARSs have been implicated in myriad dominant and recessive disease phenotypes. Glycyl‐tRNA synthetase (GARS) is a bifunctional ARS that charges tRNA^Gly in the cytoplasm and mitochondria. GARS variants have been associated with dominant Charcot‐Marie‐Tooth disease but have not been convincingly implicated in recessive phenotypes. Here, we describe a patient from the NIH Undiagnosed Diseases Program with a multisystem, developmental phenotype. Whole‐exome sequence analysis revealed that the patient is compound heterozygous for one frameshift (p.Glu83Ilefs*6) and one missense (p.Arg310Gln) GARS variant. Using in vitro and in vivo functional studies, we show that both GARS variants cause a loss‐of‐function effect: the frameshift variant results in depleted protein levels and the missense variant reduces GARS tRNA charging activity. In support of GARS variant pathogenicity, our patient shows striking phenotypic overlap with other patients having ARS‐related recessive diseases, including features associated with variants in both cytoplasmic and mitochondrial ARSs; this observation is consistent with the essential function of GARS in both cellular locations. In summary, our clinical, genetic, and functional analyses expand the phenotypic spectrum associated with GARS variants.     

KPTN gene homozygous variant‐related syndrome in the northeast of Brazil: A case report

Alteration of the KPTN gene, responsible for the coding of kaptin (a protein involved in actin cytoskeletal dynamics), causes a syndrome characterized by macrocephaly, neurodevelopmental delay and epileptic seizures. We report the first Brazilian case of KPTN gene variation, previously described in nine subjects from four interlinked families from an Amish community in Ohio, two Estonian siblings and a 9‐year‐old boy from Kansas City. We report a case of KPTN‐related syndrome in a 5‐year‐old child which presented macrocephaly, muscular hypotonia, and global development delay. The neurological examination revealed below‐expected performance in coordination and balance tests, dyspraxia, and hand‐mouth synkinesia. Expressive language was characterized by phono‐articulatory imprecision, abundance of phonological processes and morphosyntactic immaturity. Neuropsychological assessment revealed intellectual disability with impairment of verbal and executive functions. Exome sequencing was performed. Analysis revealed a homozygous 2‐nucleotide duplication c.597_598dup p.(Ser200Ilefs*55) in the KPTN gene, which is predicted to lead to a translational frameshift and formation of a premature stop codon. The phenotypic profile is similar to the cases described in the other families. Presence of macrocephaly and delayed development indicate the possibility of KPTN gene variation. Genetic testing should be carried out at an early stage in order to reach a timely diagnosis.     

Expanding the genetic and clinical spectrum of the NONO‐associated X‐linked intellectual disability syndrome

The NONO gene encodes a nuclear protein involved in RNA metabolism. Hemizygous loss‐of‐function NONO variants have been associated with syndromic intellectual disability and with left ventricular noncompaction (LVNC). A two‐year‐old boy presented to the University of Utah's Penelope Undiagnosed Disease Program with developmental delay, nonfamilial features, relative macrocephaly, and dilated cardiomyopathy with LVNC and Ebstein anomaly. Brain MRI showed a thick corpus callosum, mild Chiari I malformation, and a flattened pituitary. Exome sequencing identified a novel intronic deletion (c.154+5_154+6delGT) in the NONO gene. Splicing studies demonstrated intron 4 read‐through and the use of an alternative donor causing the frameshift p.Asn52Serfs*6. Family segregation analysis showed that the variant occurred de novo in the boy's unaffected mother. MRI and endocrine findings suggest that hypopituitarism may contribute to growth failure, abnormal thyroid hormone levels, cryptorchidism, or delayed puberty in patients with NONO‐associated disease. Also, including this case LVNC has been observed in five out of eight patients, and this report also confirms an association between loss of NONO and Ebstein anomaly. In some cases, unrelated individuals share the same pathogenic NONO variants but do not all have clinically significant LVNC, suggesting that additional modifiers may contribute to cardiac phenotypes.     

De Novo Truncating Mutations in the Kinetochore‐Microtubules Attachment Gene CHAMP1 Cause Syndromic Intellectual Disability

A rare syndromic form of intellectual disability with impaired speech was recently found associated with mutations in CHAMP1 (chromosome alignment‐maintaining phosphoprotein 1), the protein product of which is directly involved in microtubule‐kinetochore attachment. Through whole‐exome sequencing in six unrelated nonconsanguineous families having a sporadic case of intellectual disability, we identified six novel de novo truncating mutations in CHAMP1: c.1880C>G p.(Ser627*), c.1489C>T; p.(Arg497*), c.1876_1877delAG; p.(Ser626Leufs*4), c.1043G>A; p.(Trp348*), c.1002G>A; p.(Trp334*), and c.958_959delCC; p.(Pro320*). Our clinical observations confirm the phenotypic homogeneity of the syndrome, which represents therefore a distinct clinical entity. Besides, our functional studies show that CHAMP1 protein variants are delocalized from chromatin and are unable to bind to two of its direct partners, POGZ and HP1. These data suggest a pathogenic mechanism of the CHAMP1‐associated intellectual disability syndrome mediated by direct interacting partners of CHAMP1, several of which are involved in chromo/kinetochore‐related disorders.     

An Emerging Female Phenotype with Loss‐of‐Function Mutations in the Aristaless‐Related Homeodomain Transcription Factor ARX

The devastating clinical presentation of X‐linked lissencephaly with abnormal genitalia (XLAG) is invariably caused by loss‐of‐function mutations in the Aristaless‐related homeobox (ARX) gene. Mutations in this X‐chromosome gene contribute to intellectual disability (ID) with co‐morbidities including seizures and movement disorders such as dystonia in affected males. The detection of affected females with mutations in ARX is increasing. We present a family with multiple affected individuals, including two females. Two male siblings presenting with XLAG were deceased prior to full‐term gestation or within the first few weeks of life. Of the two female siblings, one presented with behavioral disturbances, mild ID, a seizure disorder, and complete agenesis of the corpus callosum (ACC), similar to the mother's phenotype. A novel insertion mutation in Exon 2 of ARX was identified, c.982delCinsTTT predicted to cause a frameshift at p.(Q328Ffs^*37). Our finding is consistent with loss‐of‐function mutations in ARX causing XLAG in hemizygous males and extends the findings of ID and seizures in heterozygous females. We review the reported phenotypes of females with mutations in ARX and highlight the importance of screening ARX in male and female patients with ID, seizures, and in particular with complete ACC.     

Maternally inherited MAF variant associated with variable expression of Aymé‐Gripp syndrome

Aymé‐Gripp syndrome is an intellectual disability syndrome characterized by autism spectrum disorder, cataracts, sensorineural hearing loss, skeletal involvement, seizures, cardiac anomalies, and distinctive facial features. The condition is caused by pathogenic variants in MAF. To date, less than 20 cases have been reported, the majority having de novo mutations. Here, we report a patient with classical features of Aymé‐Gripp syndrome who inherited a MAF variant, c.206C>G (p.P69R), from a mother with normal intellectual function and normal hearing but with cataract and significant proteinuria. To the best of our knowledge, this is the first report of a patient who inherited a MAF causative variant from a parent with normal intellect. Although the syndrome typically has multiple malformations and intellectual disability, we suggest that a mild phenotype could exist. In addition, we suggest that the basal ganglia calcifications present in our proband could be a novel finding associated with MAF variants and offer further support for the relationship between these variants and late manifestations of renal disease.     

Biallelic loss‐of‐function variants in DOCK3 cause muscle hypotonia,ataxia, and intellectual disability

DOCK3 encodes the dedicator of cytokinesis 3 protein, a member of the DOCK180 family of proteins that are characterized by guanine‐nucleotide exchange factor activity. DOCK3 is expressed exclusively in the central nervous system and plays an important role in axonal outgrowth and cytoskeleton reorganization. Dock3 knockout mice exhibit motor deficiencies with abnormal ataxic gait and impaired learning. We report 2 siblings with biallelic loss‐of‐function variants in DOCK3. Diagnostic whole‐exome sequencing (WES) and chromosomal microarray were performed on a proband with severe developmental disability, hypotonia, and ataxic gait. Testing was also performed on the proband's similarly affected brother. A paternally inherited 458 kb deletion in chromosomal region 3p21.2 disrupting the DOCK3 gene was identified in both affected siblings. WES identified a nonsense variant c.382C>G (p.Gln128*) in the DOCK3 gene (NM_004947) on the maternal allele in both siblings. Common features in both affected individuals include severe developmental disability, ataxic gait, and severe hypotonia, which recapitulates the Dock3 knockout mouse phenotype. We show that complete DOCK3 deficiency in humans leads to developmental disability with significant hypotonia and gait ataxia, probably due to abnormal axonal development.     

Biallelic mutations in mitochondrial tryptophanyl‐tRNA synthetase cause Levodopa‐responsive infantile‐onset Parkinsonism

Mitochondrial aminoacyl‐tRNA synthetases (mtARSs) are essential, ubiquitously expressed enzymes that covalently attach amino acids to their corresponding tRNA molecules during translation of mitochondrial genes. Deleterious variants in the mtARS genes cause a diverse array of phenotypes, many of which involve the nervous system. Moreover, distinct mutations in mtARSs often cause different clinical manifestations. Recently, the gene encoding mitochondrial tryptophanyl tRNA synthetase (WARS2) was reported to cause 2 different neurological phenotypes, a form of autosomal recessive intellectual disability and a syndrome of severe infantile‐onset leukoencephalopathy. Here, we report the case of a 17‐year‐old boy with compound heterozygous mutations in WARS2 (p.Trp13Gly, p.Ser228Trp) who presented with infantile‐onset, Levodopa‐responsive Parkinsonism at the age of 2 years. Analysis of patient‐derived dermal fibroblasts revealed decreased steady‐state WARS2 protein and normal OXPHOS content. Muscle mitochondrial studies suggested mitochondrial proliferation without obvious respiratory chain deficiencies at the age of 9 years. This case expands the phenotypic spectrum of WARS2 deficiency and emphasizes the importance of mitochondrial protein synthesis in the pathogenesis of Parkinsonism.     

Mutations of the aminoacyl‐tRNA‐synthetases SARS and WARS2 are implicated in the etiology of autosomal recessive intellectual disability

Intellectual disability (ID) is the hallmark of an extremely heterogeneous group of disorders that comprises a wide variety of syndromic and non‐syndromic phenotypes. Here, we report on mutations in two aminoacyl‐tRNA synthetases that are associated with ID in two unrelated Iranian families. In the first family, we identified a homozygous missense mutation (c.514G>A, p.Asp172Asn) in the cytoplasmic seryl‐tRNA synthetase (SARS) gene. The mutation affects the enzymatic core domain of the protein and impairs its enzymatic activity, probably leading to reduced cytoplasmic tRNA^Ser concentrations. The mutant protein was predicted to be unstable, which could be substantiated by investigating ectopic mutant SARS in transfected HEK293T cells. In the second family, we found a compound heterozygous genotype of the mitochondrial tryptophanyl‐tRNA synthetase (WARS2) gene, comprising a nonsense mutation (c.325delA, p.Ser109Alafs*15), which very likely entails nonsense‐mediated mRNA decay and a missense mutation (c.37T>G, p.Trp13Gly). The latter affects the mitochondrial localization signal of WARS2, causing protein mislocalization. Including AIMP1, which we have recently implicated in the etiology of ID, three genes with a role in tRNA‐aminoacylation are now associated with this condition. We therefore suggest that the functional integrity of tRNAs in general is an important factor in the development and maintenance of human cognitive functions.     

The RBMX gene as a candidate for the Shashi X‐linked intellectual disability syndrome

A novel X‐linked intellectual disability (XLID) syndrome with moderate intellectual disability and distinguishing craniofacial dysmorphisms had been previously mapped to the Xq26‐q27 interval. On whole exome sequencing in the large family originally reported with this disorder, we identified a 23 bp frameshift deletion in the RNA binding motif protein X‐linked (RBMX) gene at Xq26 in the affected males (n = 7), one carrier female, absent in unaffected males (n = 2) and in control databases (7800 exomes). The RBMX gene has not been previously causal of human disease. We examined the genic intolerance scores for the coding regions and the non‐coding regions of RBMX; the findings were indicative of RBMX being relatively intolerant to loss of function variants, a distinctive pattern seen in a subset of XLID genes. Prior expression and animal modeling studies indicate that loss of function of RBMX results in abnormal brain development. Our finding putatively adds a novel gene to the loci associated with XLID and may enable the identification of other individuals affected with this distinctive syndrome.     

De novo loss‐of‐function mutations in X‐linked SMC1A cause severe ID and therapy‐resistant epilepsy in females: expanding the phenotypic spectrum

De novo missense mutations and in‐frame coding deletions in the X‐linked gene SMC1A (structural maintenance of chromosomes 1A), encoding part of the cohesin complex, are known to cause Cornelia de Lange syndrome in both males and females. For a long time, loss‐of‐function (LoF) mutations in SMC1A were considered incompatible with life, as such mutations had not been reported in neither male nor female patients. However, recently, the authors and others reported LoF mutations in females with intellectual disability (ID) and epilepsy. Here we present the detailed phenotype of two females with de novo LoF mutations in SMC1A, including a de novo mutation of single base deletion [c.2364del, p.(Asn788Lysfs*10)], predicted to result in a frameshift, and a de novo deletion of exon 16, resulting in an out‐of‐frame mRNA splice product [p.(Leu808Argfs*6)]. By combining our patients with the other recently reported females carrying SMC1A LoF mutations, we ascertained a phenotypic spectrum of (severe) ID, therapy‐resistant epilepsy, absence/delay of speech, hypotonia and small hands and feet. Our data show the existence of a novel phenotypic entity – distinct from CdLS – and caused by de novo SMC1A LoF mutations.     

A loss‐of‐function homozygous mutation in DDX59 implicates a conserved DEAD‐box RNA helicase in nervous system development and function

We report on a homozygous frameshift deletion in DDX59 (c.185del: p.Phe62fs*13) in a family presenting with orofaciodigital syndrome phenotype associated with a broad neurological involvement characterized by microcephaly, intellectual disability, epilepsy, and white matter signal abnormalities associated with cortical and subcortical ischemic events. DDX59 encodes a DEAD‐box RNA helicase and its role in brain function and neurological diseases is unclear. We showed a reduction of mutant cDNA and perturbation of SHH signaling from patient‐derived cell lines; furthermore, analysis of human brain gene expression provides evidence that DDX59 is enriched in oligodendrocytes and might act within pathways of leukoencephalopathies‐associated genes. We also characterized the neuronal phenotype of the Drosophila model using mutant mahe, the homolog of human DDX59, and showed that mahe loss‐of‐function mutant embryos exhibit impaired development of peripheral and central nervous system. Taken together, our results support a conserved role of this DEAD‐box RNA helicase in neurological function.     

De novo heterozygous missense and loss‐of‐function variants in CDC42BPB are associated with a neurodevelopmental phenotype

CDC42BPB encodes MRCKβ (myotonic dystrophy‐related Cdc42‐binding kinase beta), a serine/threonine protein kinase, and a downstream effector of CDC42, which has recently been associated with Takenouchi‐Kosaki syndrome, an autosomal dominant neurodevelopmental disorder. We identified 12 heterozygous predicted deleterious variants in CDC42BPB (9 missense, 2 frameshift, and 1 nonsense) in 14 unrelated individuals (confirmed de novo in 11/14) with neurodevelopmental disorders including developmental delay/intellectual disability, autism, hypotonia, and structural brain abnormalities including cerebellar vermis hypoplasia and agenesis/hypoplasia of the corpus callosum. The frameshift and nonsense variants in CDC42BPB are expected to be gene‐disrupting and lead to haploinsufficiency via nonsense‐mediated decay. All missense variants are located in highly conserved and functionally important protein domains/regions: 3 are found in the protein kinase domain, 2 are in the citron homology domain, and 4 in a 20‐amino acid sequence between 2 coiled‐coil regions, 2 of which are recurrent. Future studies will help to delineate the natural history and to elucidate the underlying biological mechanisms of the missense variants leading to the neurodevelopmental and behavioral phenotypes.     

Deficient activity of alanyl‐tRNA synthetase underlies an autosomal recessive syndrome of progressive microcephaly,hypomyelination, and epileptic encephalopathy

Aminoacyl‐transfer RNA (tRNA) synthetases ligate amino acids to specific tRNAs and are essential for protein synthesis. Although alanyl‐tRNA synthetase (AARS) is a synthetase implicated in a wide range of neurological disorders from Charcot‐Marie‐Tooth disease to infantile epileptic encephalopathy, there have been limited data on their pathogenesis. Here, we report loss‐of‐function mutations in AARS in two siblings with progressive microcephaly with hypomyelination, intractable epilepsy, and spasticity. Whole‐exome sequencing identified that the affected individuals were compound heterozygous for mutations in AARS gene, c.2067dupC (p.Tyr690Leufs*3) and c.2738G>A (p.Gly913Asp). A lymphoblastoid cell line developed from one of the affected individuals showed a strong reduction in AARS abundance. The mutations decrease aminoacylation efficiency by 70%–90%. The p.Tyr690Leufs*3 mutation also abolished editing activity required for hydrolyzing misacylated tRNAs, thereby increasing errors during aminoacylation. Our study has extended potential mechanisms underlying AARS‐related disorders to include destabilization of the protein, aminoacylation dysfunction, and defective editing activity.     

Two Siblings with Homozygous Pathogenic Splice‐Site Variant in Mitochondrial Asparaginyl–tRNA Synthetase (NARS2)

A homozygous missense mutation (c.822G>C) was found in the gene encoding the mitochondrial asparaginyl–tRNA synthetase (NARS2) in two siblings born to consanguineous parents. These siblings presented with different phenotypes: one had mild intellectual disability and epilepsy in childhood, whereas the other had severe myopathy. Biochemical analysis of the oxidative phosphorylation (OXPHOS) complexes in both siblings revealed a combined complex I and IV deficiency in skeletal muscle. In‐gel activity staining after blue native‐polyacrylamide gel electrophoresis confirmed the decreased activity of complex I and IV, and, in addition, showed the presence of complex V subcomplexes. Considering the consanguineous descent, homozygosity mapping and whole‐exome sequencing were combined revealing the presence of one single missense mutation in the shared homozygous region. The c.822G>C variant affects the 3′ splice site of exon 7, leading to skipping of the whole exon 7 and a part of exon 8 in the NARS2 mRNA. In EBV‐transformed lymphoblasts, a specific decrease in the amount of charged mt‐tRNA^Asn was demonstrated as compared with controls. This confirmed the pathogenic nature of the variant. To conclude, the reported variant in NARS2 results in a combined OXPHOS complex deficiency involving complex I and IV, making NARS2 a new member of disease‐associated aaRS2.     

Tatton‐Brown‐Rahman syndrome: Six individuals with novel features

Tatton‐Brown Rahman syndrome (TBRS) is an overgrowth‐intellectual disability syndrome caused by heterozygous variants in DNMT3A. Seventy‐eight individuals have been reported with a consistent phenotype of somatic overgrowth, mild to moderate intellectual disability, and similar dysmorphisms. We present six individuals with TBRS, including the youngest individual thus far reported, first individual to be diagnosed with tumor testing and two individuals with variants at the Arg882 domain, bringing the total number of reported cases to 82. Patients reported herein have additional clinical features not previously reported in TBRS. One patient had congenital diaphragmatic hernia. One patient carrying the recurrent p.Arg882His DNMT3A variant, who was previously reported as having a phenotype due to a truncating variant in the CLTC gene, developed a ganglioneuroblastoma at 18 months and T‐cell lymphoblastic lymphoma at 6 years of age. Four patients manifested symptoms suggestive of autonomic dysfunction, including central sleep apnea, postural orthostatic hypotension, and episodic vasomotor instability in the extremities. We discuss the molecular and clinical findings in our patients with TBRS in context of existing literature.